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Fishery Bulletin 90(2|. 1992 



were assayed in each tissue using 

 a variety of electrophoresis buffers. 

 On the basis of the aforemen- 

 tioned analyses, three tissues 

 were retained from all conch col- 

 lected subsequently in 1988-90: 

 (1) foot muscle, (2) proboscis with 

 radula, and (3) digestive gland 

 with gonad. The three tissues 

 were dissected from each individ- 

 ual, placed in separate cryotubes 

 or plastic bags, and frozen in the 

 field with liquid nitrogen or dry 

 ice. All tissues were stored at 

 -80°C until prepared for en- 

 zyme extraction. 



Electrophoresis 



Allozymes were detected by hori- 

 zontal starch-gel electrophore- 

 sis following the procedures of 

 Aebersold et al. (1987). Enzymes 

 were extracted by homogenizing 

 each tissue separately in 0.5-1.0 

 volumes of 0.05M PIPES, 0.05% 

 Triton X-100, and 0.2 mM pyri- 

 doxal-5'-phosphate (adjusted to 

 pH 6.8 with l.OM NaOH). Gels 

 were prepared with a 12.5% mix- 

 ture (wt:vol) of Connaught starch 

 (Fisher Sci. Co.) and one of five 

 buffer solutions (Table 2). Histo- 

 chemical staining of gels followed 

 standard procedures (Morizot and Schmidt 1990). Gels 

 were stained by agar overlay for all enzymes except 

 AAT. 



Presumptive loci and alleles were designated by the 

 nomenclature system outlined by Shaklee et al. (1990), 

 except peptidase loci were identified by their di- or tri- 

 peptide acronyms (DPEP, TPEP). Multiple loci of a 

 particular enzyme were designated numerically (1,2, 

 etc.) from fastest to slowest anodic mobility. Alleles of 

 a particular locus were designated by their relative, 

 anodic mobilities (most frequent allele = *100). 



Statistics 



Genotypic proportions at each locus were tested for 

 goodness-of-fit to Hardy-Weinberg expectations using 

 the likelihood-ratio test or G -statistic (Sokal and Rohlf 

 1981). Allele frequencies at each locus were tested for 

 homogeneity among samples by contingency table 

 (samples x alleles) G -tests (Sokal and Rohlf 1981). This 

 total G -statistic, or likelihood ratio, was then parti- 



tioned into hierarchical components representing 

 temporal and spatial components of genetic variation 

 within and among localities, respectively (e.g., Smouse 

 and Ward 1978). An approximate F -ratio was then con- 

 structed as (G among localities/df)/(G among years 

 within localities/df) to test whether the genetic hetero- 

 geneity among localities was significantly greater than 

 the heterogeneity among years within localities. The 

 total gene diversity (Nei 1973) was similarly partitioned 

 into within- and among-locality components following 

 the algorithm of Chakraborty et al. (1982). In all tests 

 of statistical significance, significance probabilities 

 were adjusted for the number of tests (loci) evaluated 

 simultaneously (Rice 1989). 



Nei's (1972) index of gene identity was calculated be- 

 tween all population samples. The genetic similarities 

 among all populations, including those sampled by Mit- 

 ton et al. (1989), were represented graphically in a 

 UPGMA dendrogram (Sneath and Sokal 1973). The 

 1987 sample from Ballast Key was excluded from these 

 latter analyses because of small sample size (n 12). 



